Protective Effects Of Rhubarb On Septic Rats’ Brain Function

Rhubarb is called General because of its rapid and strong effects,which is famous for the pharmacological action of diarrhea,correlated with water adjustment of colon.Recently,more and more researches reveal a kind of protein,which is called AQP,have closely relationship with water absorption and secretion.They found restrained water absorption in colon could led to diarrhea,and to dieresis in kidney.Futhermore,the diarrhea and dieresis effects of Rhubarb are correlated with aquaporins in the both organs.Then,we will try to find out whether AQPs participates in the formation of cerebral edema after sepsis,and Rhubarb could alleviate the procedure.Meanwhile we want to investigate whether the pathogenesis of SAE is the exaggerated inflammatory response,and the later can lead to irreversible damage of brain parenchyma cells.Previous studies showed that rhubarb could reduce the incidence and mortality of multiple organ dysfunction syndrome(MODS)because of its anti-inflammatory activities.We are eager to know whether the traditional chinese medicine,rhubarb,could prevent the negative effects of inflammatory factors on the septic rats’ brain tissue.Numerous researches demonstrated that the relationship of the increased permeability of blood brain barrier and sepsis associated encephalopathy(SAE)is quite closely.As we all know,one mechanism of the major brain edema is the increased permeability of blood brain barrier.Researchers found rhubarb could decrease BBB permeability.It would be worthy to find out the role of rhubarb in BBB permeability after sepsis.According to the above problems,the study include two parts: Part I : ffects of Rhubarb on brain dysfunction in septic ratsObjective To investigate the effect of Rhubarb on brain dysfunction in septic rats.Methods 84 Wister male rats were randomly divided into three groups(n=28 each),sham operation group(Sham),cecal ligation and puncture group(CLP),Rhubarb+ CLP group(R+CLP).every groups were divided into two subgroups(A and B).Female rats were gavaged daily(1ml/100 g boody weight)with Rhubarb for three days.The same dose of saline was used in the control groups.Models of rats with sepsis were established by CLP.Survival rate was monitored in A groups,10 days totally.In B groups,animals were euthanized at 12、24 and 48 hours after CLP.Wet-dry weighting method was used to detect the brain water content(BWC).The ELISA method was employed to determine the concentrations of plasma S100β,Neuron-specific enolase(NSE)and the expression levels of brain tissue NF-κB,IL-6 and TNF-αproteins.Then Hematoxylin and eosin(HE)staining was used to detect brain pathologic change.Immunohistochemical SP method was applied to count the number of AQP-4 positive glial cells in brain tissues.Results The BWC values of the CLP and R+CLP group at all time points were significantly higher than those of the sham group at the same periods(P <0.05),and which in R+CLP group was lower than that in CLP group at 6h after CLP(P <0.05).CLP and R+CLP groups brain tissue levels of NF-ΚB,IL-6 and TNF-ɑ were significantly increased compared with Sham group at each moment after CLP(P<0.05),and R+CLP group brain tissue levels of NF-ΚB,IL-6 and TNF-ɑ were also significantly lower than the CLP group at 24 h after CLP(P<0.05).Compared with Sham group,the concentrations of plasma S100β,NSE were notably increased after CLP(P<0.05)at each moment,and which were significantly reversed by treatment with Rhubarb at 24 h after CLP(P<0.05).Brain tissue HE staining showed the pathological damage were lightest in the sham group,However,the pathological damage in the CLP group was the most serious degree,and compared with the CLP group,the pathological damage was significantly decreased in the R+CLP group at 6h after CLP(P<0.05).The AQP-4 positive gliacyte cells in brain tissues in the CLP group were more than those in the sham group at each time(P<0.05),But those were significantly fewer than those in the CLP group at 6h after CLP(P<0.05).There was a significantly positive correlation between AQP-4 expression and brain water content(r = 0.895,P<0.05).Rhubarb could prevented mortality in septic rats.Conclusions Rhubarb can inhibit the expressions of IL-6,and TNF-ɑ proteins in rats with sepsis by inhibit NF-κB signaling pathway,and ameliorated the CLP-induced brain tissue edema by down-regulating the expression of the brain gliacyte AQP-4.Part II: Effects of Rhubarb on Blood Brain Barrier in Septic RatsObjective To investigate the effect of Rhubarb on blood brain barrier in septic rats.Methods 24 Wister rats were randomly divided into two groups(n=12 each).controlled(A)group,experimental(B)group.Experimental rats were gavaged daily(1ml/100 g body weight)with Rhubarb for three days.The same dose of solvent was used in the controlled groups.Models of rats with sepsis were established by CLP.The changes of their neurological behaviors were observed.0.5% Evans Blue(EB)was injected through caudal vein(12ml·kg-1,i.v.)before 0.5h of sacrificing the animals.Then,normal saline were used to remove the intravascular dye.Brains were removed euthanized at 24 hours after CLP.BBB permeability by EB method.the content of MMP-2、MMP-9、TIMP and collagen protein-Ⅳ were detected by Elisa.Results The value of neurological behaviors in the experimental group is significantly incresed compared with controlled group at at 24 h after CLP.In contrast to the controlled group,the concentration of EB were decreased at 24 hours in experimental group after CLP.The content of MMP-2 and MMP-9 in brain tissues in the experimental group were less than those in the controlled group at 24 h,wheras,TIMP and collagen protein-Ⅳ in the former group were more than those in the latter group at 24 h.There was a significantly positive correlation between MMP-2、MMP-9、TIMP and collagen protein-Ⅳ.Conclusions Rhubarb can ameliorate the CLP-induced brain tissue edema and BBB permeability by down-regulating the expression of the brain MMP-2 and MMP-9,up-regulating the expression of TIMP and collagen protein-Ⅳ.